Abstract: :
Purpose: Nonmuscle myosin II proteins, the conventional formof myosin present in most eukaryotic cells, are active in thecontractile process during embryonic development as well aslater life. These proteins contribute to the mechanisms by whichcells migrate, change their shape and undergo cytokinesis. Tostudy the in vivo function of the three isoforms of the nonmusclemyosin heavy chain (NMHC), II–A, II–B, and II–C,we have generated mice that have been ablated for the NMHC II–Bgene product, as well as mice harboring a single amino acidmutation in NMHC II–B. This mutation, in which Arg709was replaced by Cys (R709C) was introduced to produce a putativemouse model for human retinal disease. Previous work from thislaboratory showed that ablation of NMHC II–B resultedin rosette formation in the retina just prior to birth (J CompNeurol 433:62–74. ) Here we report on the C/C and R/Cmice.Methods: Transgenic mice were generated by homologous recombination.The point mutation R709C was introduced into exon 17. Aftergeneration of heterozygous ES cells the mutation was confirmedby nucleotide sequencing and positive ES cell clones were injectedinto blastocysts from C57BJ/6 mice. Eyes were enucleated fromnewborn myosin–IIB knockout mice, and from adult R/R andR/C knockin mice. Embryos were obtained at day E14 from knockout,R/R, and C/C knockin mice as well as from adult R/C mice. Allspecimens were formalin–fixed and paraffin embedded forroutine histology. Immunohistochemistry was performed usingantibodies to nonmuscle myosin–IIA, –IIB, –IIC,GFAP and NSE.Results: Eyes taken from embryos of R/R and C/C mice showedno morphological differences; however C/C mice die at E14.5due to cardiac and brain defects, preventing further study.The retinas from adult R/C mice showed an increase in the numberof nuclei in the inner plexiform layer as well as retarded migrationof the ganglion cells. This suggests a possible arrest in retinalcell migration. Immunohistochemistry demonstrated similar levelsand patterns of staining for nonmuscle myosin–IIA on vesselsand a low expression of nonmuscle myosin–IIC in the retina.Conclusions: Adult R/C mice with a single amino acid mutationin NMHC II–B show abnormalities in neuronal cell migrationin the adult retina. The significance of these findings hasyet to be determined, but since it is likely that, similar tomice, only humans that are heterozygous for this mutation willsurvive, it will be of interest to see if humans bearing a mutationin NMHC II–B manifest similar abnormalities in their retinas.